Abstract

Melanoma is known to be highly resistant to chemotherapy. Treatment with high dose IL-2 has shown significant clinical benefit in a minority of metastatic melanoma patients and has lead to long term survival in a few cases. However, this treatment is associated with excessive multiorgan toxicities which severely limit its use. We propose that one mechanism of effective IL-2 therapy is through the direct activation of IL-24 on melanoma tumors and subsequent IL-24 mediated tumor growth suppression or apoptosis. To test this proposed mechanism, five melanoma cell lines were treated with increasing amounts of recombinant hIL-2 ranging from 250 to 2000 U/ml. Three out of five cell lines (A375, WM1341, WM793) showed increasing levels of IL-24 protein when measured by Western blotting, while the remaining two lines (WM35, MeWo) remained negative for IL-24. A375, a highly metastatic cell line, showed an average increase in IL-24 protein of 43% (p=0.03) when cells were incubated 1000 U/ml of IL-2 for 24 hr, while IL-24 protein in WM1341 treated with an equal concentration of IL-2 increased by a statistically significant 31.6% (p=0.0029) over basal levels. This increase in IL-24 was abolished by either preincubating with an anti-IL-2 antibody or by neutralizing the IL-2 receptor directly with antibodies against the receptor chains. These cell lines were found to contain IL-2R\#946; message by RT-PCR. Thus we believe that IL-2 is signaling through IL-2R\#946; on some melanoma cells to upregulate IL-24 protein expression. To address the function of IL-2 in melanoma cell lines, we treated A375 with 1000 U/ml of IL-2 for six days. Cell viability was determined and found to be significantly decreased by 18% compared to untreated control cells (p=0.01). We also demonstrated by ELISA that these cells secrete IL-24 protein in higher amounts when stimulated with IL-2 than control cells. These data support our hypothesis that in addition to its immunotherapeutic effects, IL-2 also acts directly on some melanoma tumors. Experiments are in progress to determine the mechanism of IL-2 dependent upregulation of IL-24.